Recent clinical trials (e.g., CAST and CAST-II) established a cardiotoxic effect of treatment of ventricular arrhythmias with Class I antiarrhythmic agents (Na+ channel blockers). This failure has shifted clinical interest to Class III antiarrhythmic agents (K+ channel blockers). However, antiarrhythmic drugs that block K+ channels vary in selectivity and display conformation specific interactions. Such conformation specific interactions can cause the degree of block of a channel to vary by orders of magnitude depending on the rate, duration and potentials to which the channel is subject. Such conformation dependent binding can be either detrimental or beneficial. This proposal focuses on the relationship between antiarrhythmic drug binding and a particular class of conformational changes, namely C-type inactivation in cardiac K+ channels. C-type inactivation is more widely distributed among cardiac K+ channels than N-type and may be the dominant determinant of such important properties as recovery, K+ sensitivity and drug use-dependent. The goal of this proposal is to elucidate how C-type inactivation can influence the complex patterns of block and use-dependence seen with Class III agents. The central hypothesis is that C-type inactivation involves two conformational changes, closure of the external pore mouth, and closure of the internal pore mouth. Closure of the internal mouth of the channel is proposed to occur through a lipophilic collapse involving the S6 domain. Intracellular lipophilic drug binding promotes this collapse by excluding K+ and increasing the net lipophilic environment. The PI further hypothesizes that the conformational changes in the two regions are not independent but are coupled. The proposed physical and energetic mechanisms will be simulated and tested experimentally to provide a quantitative model of C-type inactivation and antiarrhythmic drug binding. This study will provide a molecular basis for the use dependent properties of a broad class